In order to gain insight into the mechanisms of insulin-like growth factor I (IGF I) signaling in cardiac muscle, the applicant plans to study the initial, intermediate and distal IGF I signaling steps that are involved in the regulation of myocardial growth and function. Among the distal signaling pathways of IGF I, c-fos and c-jun play important roles in the regulation of myocardial growth and function. In addition, preliminary studies indicate that p53, a key regulator of cell cycle progression, is a novel element of IGF I signaling in cardiac muscle cells. The hypothesis to be tested is that IGF I actions on cardiac muscle involve c-fos/c-jun pathways and p53 pathway. The specific aims are: 1) to define in vivo IGF I effects on initial signaling events, i.e., on the phosphorylation of IGF I receptor and the endogenous substrate IRS I; 2) to define the in vivo effects of IGF I on intermediate steps of IGF I signaling pathways in cardiac muscle, including the activation of phosphatidylinositol 3-kinase (PI 3 kinase) and the tyrosine phosphorylation and activation of mitogen-activated protein (MAP kinase); 3) to study the distal IGF I signaling events by: a) studying the effects of IGF I on the expression and phosphorylation of c-fos and c-jun in cardiac muscle in intact rat myocardium and cultured cardiac muscle cells; b) investigating the actions of IGF I on the cis-acting elements of c-fos and c-jun in cardiac muscle cells in cultured cardiac muscle cells transfected with the TPA responsive element (TRE) or the serum response element, as well as determine the binding of specific nuclear proteins to the SRE cis-acting elements; c) investigating the regulation of p53 expression and phosphorylation by IGF I in cardiac muscle in vivo and in cultured cardiac muscle cells; d) defining the regulatory effects of IGF I on the binding of p53 to its cis-acting element and on the activation of WAF1 transcription in cardiac muscle cells; 4) to determine whether IGF I effects on c-fos/c-jun and p53 pathways require activation of PI 3 kinase and MAP kinases in cardiac muscle; 5) to investigate the regulatory effects of IGF I on the progression of cell cycle in cardiac muscle cells by double-labeling cells with propidium iodine and bromodeoxyuridine.